scholarly journals The hydrolysis of α-human atrial natriuretic peptide by pig kidney microvillar membranes is initiated by endopeptidase-24.11

1987 ◽  
Vol 243 (1) ◽  
pp. 183-187 ◽  
Author(s):  
S L Stephenson ◽  
A J Kenny
Keyword(s):  
Atrial Natriuretic Peptide ◽  
Natriuretic Peptide ◽  
Angiotensin I ◽  
Human Atrial Natriuretic Peptide ◽  
Pig Kidney ◽  
Angiotensin I Converting Enzyme ◽  
Endopeptidase 24.11 ◽  
Hydrolysis Of ◽  
Atrial Natriuretic

alpha-Human atrial natriuretic peptide, a 28-amino-acid-residue peptide, was rapidly hydrolysed by pig kidney microvillar membranes in vitro, with a t1/2 of 8 min, comparable with the rate observed with angiotensins II and III. The products of hydrolysis were analysed by h.p.l.c., the pattern obtained with membranes being similar to that with purified endopeptidase-24.11 (EC 3.4.24.11). No hydrolysis by peptidyl dipeptidase A (angiotensin I converting enzyme, EC 3.4.15.1) was observed. The contribution of the various microvillar membrane peptidases was assessed by including specific inhibitors. Phosphoramidon, an inhibitor of endopeptidase-24.11, caused 80-100% suppression of the products. Captopril and amastatin (inhibitors of peptidyl dipeptidase A and aminopeptidases respectively) had no significant effect. Hydrolysis at an undefined site within the disulphide-linked ring occurred rapidly, followed by hydrolysis at other sites, including the Ser25--Phe26 bond.

scholarly journals Hydrolysis of α-human atrial natriuretic peptide in vitro by human kidney membranes and purified endopeptidase-24.11. Evidence for a novel cleavage site

1988 ◽  
Vol 254 (2) ◽  
pp. 531-537 ◽  
Author(s):  
Y Vanneste ◽  
A Michel ◽  
R Dimaline ◽  
T Najdovski ◽  
M Deschodt-Lanckman
Keyword(s):  
Atrial Natriuretic Peptide ◽  
Natriuretic Peptide ◽  
Bond Cleavage ◽  
Human Kidney ◽  
Human Atrial Natriuretic Peptide ◽  
Endopeptidase 24.11 ◽  
Therapeutic Uses ◽  
Atrial Natriuretic

alpha-Human atrial natriuretic peptide (hANP) is secreted by the heart and acts on the kidney to promote a strong diuresis and natriuresis. In vivo it has been shown to be catabolized partly by the kidney. Crude microvillar membranes of human kidney degrade 125I-ANP at several internal bonds generating metabolites among which the C-terminal fragments were identified. Formation of the C-terminal tripeptide was blocked by phosphoramidon, indicating the involvement of endopeptidase-24.11 in this cleavage. Subsequent cleavages by aminopeptidase(s) yielded the C-terminal dipeptide and free tyrosine. Using purified endopeptidase 24.11, we identified seven sites of hydrolysis in unlabelled alpha-hANP: the bonds Arg-4-Ser-5, Cys-7-Phe-8, Arg-11-Met-12, Arg-14-Ile-15, Gly-16-Ala-17, Gly-20-Leu-21 and Ser-25-Phe-26. However, the bonds Gly-16-Ala-17 and Arg-4-Ser-5 did not fulfil the known specificity requirements of the enzyme. Cleavage at the Gly-16-Ala-17 bond was previously observed by Stephenson & Kenny [(1987) Biochem. J. 243, 183-187], but this is the first report of an Arg-Ser bond cleavage by this enzyme. Initial attack of alpha-hANP by endopeptidase-24.11 took place at a bond within the disulphide-linked loop and produced a peptide having the same amino acid composition as intact ANP. The bond cleaved in this metabolite was determined as the Cys-7-Phe-8 bond. Determination of all the bonds cleaved in alpha-hANP by endopeptidase-24.11 should prove useful for the design of more stable analogues, which could have therapeutic uses in hypertension.

Direct effect of α-human atrial natriuretic peptide on human vasculature in vivo and in vitro

1988 ◽  
Vol 74 (2) ◽  
pp. 207-211 ◽  
Author(s):  
A. Hughes ◽  
S. Thom ◽  
P. Goldberg ◽  
G. Martin ◽  
P. Sever
Keyword(s):  
Blood Flow ◽  
Atrial Natriuretic Peptide ◽  
Natriuretic Peptide ◽  
Forearm Blood Flow ◽  
Renin Angiotensin System ◽  
Organ Bath ◽  
Human Atrial Natriuretic Peptide ◽  
Atrial Natriuretic

1. The effect of a α-human atrial natriuretic peptide (1–28) (ANP) on human vasculature was investigated in vivo and in vitro. Possible involvement of vascular dopamine receptors and the renin-angiotensin system in the response to ANP was also studied in vivo. 2. Forearm blood blow was measured by venous occlusion plethysmography. Isolated human blood vessels were studied using conventional organ bath techniques. 3. ANP (0.1–1 μg/min, intra-arterially) produced a dose-dependent increase in forearm blood flow, corresponding to a 163% increase in net forearm blood flow in the study arm. This action of ANP was not antagonized by (R)-sulpiride (100 μg/min, intra-arterially), a selective vascular dopamine receptor antagonist, or 50 mg of oral captopril, an inhibitor of angiotensin-converting enzyme. 4. ANP (1 nmol/l–1 μmol/l) produced concentration-dependent relaxation of isolated human arteries, including brachial artery, but was without effect on isolated human saphenous vein. 5. ANP produces vasodilatation in vivo and relaxes isolated human arterial smooth muscle. This action of ANP may contribute to its reported hypotensive effects in vivo.

scholarly journals Metabolism of neuropeptides. Hydrolysis of the angiotensins, bradykinin, substance P and oxytocin by pig kidney microvillar membranes

1987 ◽  
Vol 241 (1) ◽  
pp. 237-247 ◽  
Author(s):  
S L Stephenson ◽  
A J Kenny
Keyword(s):  
Substance P ◽  
Membrane Fraction ◽  
Angiotensin I ◽  
Peptide Bonds ◽  
Prolonged Incubation ◽  
Pig Kidney ◽  
Angiotensin I Converting Enzyme ◽  
Endopeptidase 24.11 ◽  
20 Nm ◽  
Hydrolysis Of

Microvillar membranes derived from the brush border of the renal proximal tubule are very rich in peptidases. Pig kidney microvilli contain endopeptidase-24.11 associated with a battery of exopeptidases. The manner by which some neuropeptides are degraded by the combined attack of the peptidases of this membrane has been investigated. The contribution of individual peptidases was assessed by including inhibitors (phosphoramidon, captopril, amastatin and di-isopropyl fluorophosphate) with the membrane fraction when incubated with the peptides. Substance P, bradykinin and angiotensins I, II and III and insulin B-chain were rapidly hydrolysed by kidney microvilli. Oxytocin was hydrolysed much more slowly, but no products were detected from [Arg8]vasopressin or insulin under the conditions used for other peptides. The peptide bonds hydrolysed were identified and the contributions of the different peptidases were quantified. For each of the susceptible peptides, the main contribution came from endopeptidase-24.11 (inhibited by phosphoramidon). Peptidyl dipeptidase A (angiotensin-I-converting enzyme) was of less importance, even in respect of angiotensin I and bradykinin. When [2,3-Pro3,4-3H]bradykinin was also investigated at a lower concentration (20 nM), the conclusions in regard to the contributions of the two peptidases were unchanged. The possibility that endopeptidase-24.11 might attack within the six-residue disulphide-bridged rings of oxytocin and vasopressin was examined by dansyl(5-dimethylaminonaphthalene-1-sulphonyl)ation and by reduction and carboxymethylation of the products after incubation. Additional peptides were only observed after prolonged incubation, consistent with hydrolysis at the Tyr2-Ile3 and Tyr2-Phe3 bonds respectively. These results show that a range of neuropeptides are efficiently degraded by microvillar membranes and that endopeptidase-24.11 plays a key role in this process.

In vitro stability of human atrial natriuretic peptide(h-ANP)

1991 ◽  
Vol 203 (2-3) ◽  
pp. 235-241 ◽  
Author(s):  
Erika Artner-Dworzak ◽  
Herbert Lindner ◽  
Bernd Puschendorf
Keyword(s):  
Atrial Natriuretic Peptide ◽  
Natriuretic Peptide ◽  
Human Atrial Natriuretic Peptide ◽  
In Vitro Stability ◽  
Atrial Natriuretic

scholarly journals Hydrolysis of human and pig brain natriuretic peptides, urodilatin, C-type natriuretic peptide and some C-receptor ligands by endopeptidase-24.11

1993 ◽  
Vol 291 (1) ◽  
pp. 83-88 ◽  
Author(s):  
A J Kenny ◽  
A Bourne ◽  
J Ingram
Keyword(s):  
Atrial Natriuretic Peptide ◽  
Brain Natriuretic Peptide ◽  
Natriuretic Peptide ◽  
Natriuretic Peptides ◽  
Receptor Ligands ◽  
Prolonged Incubation ◽  
Endopeptidase 24.11 ◽  
Pig Brain ◽  
Hydrolysis Of ◽  
Atrial Natriuretic

Endopeptidase-24.11 (E-24.11, EC 3.4.24.11) is widely believed to play a physiological role in metabolizing atrial natriuretic peptide (ANP). Since the discovery of ANP, new natriuretic peptides have been isolated and other peptides synthesized as receptor ligands. The hydrolysis in vitro of six related peptides by the endopeptidase has been studied, mainly by h.p.l.c. The initial attack on the 32-residue form of pig brain natriuretic peptide (pBNP-32) was shown to be at the Ser20-Leu21 bond, as had been previously shown for the 26-residue form. In contrast, human brain natriuretic peptide-32 (hBNP-32), which differs in ten residues from pBNP-32, was attacked first at the Met4-Val5 bond, releasing the N-terminal tetrapeptide, and only later at bonds within the ring: at Arg17-Ile18 and subsequently at four other sites. Urodilatin, which has a four-residue extension at the N-terminus compared with alpha-human atrial natriuretic peptide-28 (alpha-hANP), was degraded at about half the rate of the latter, though the C-terminal Phe-Arg-Tyr was released at the same rate. The 22-residue C-type natriuretic peptide was hydrolysed more rapidly than alpha-hANP, as were two C-receptor ligands (peptides with deletions within the ring): C-ANP4-23 (rANP4-23 des-Gln18,Ser19,Gly20,Leu21,Gly22) and SC 46542 (hANP5-28 des-Phe8,Gly9,Ala17,Gln18). Angiotensin-converting enzyme failed to hydrolyse pBNP-32, hBNP-32 or 125I-rat (r) ANP, even after prolonged incubation. Km and kcat values were determined for the hydrolysis of alpha-hANP, porcine BNP-26, porcine BNP-32 and 125I-rANP by E-24.11. Ki values were determined for six peptides, alpha-hANP, urodilatin, hBNP-32, C-type natriuretic peptide (CNP), SC 46542 and C-type natriuretic peptide (C-ANP4-23), in radiometric assays of E-24.11 with either [125I] insulin B chain or [125I] rANP as substrate. The Ki values (2.5-13 microM) for CNP were the lowest of any of the group, whereas those for hBNP-32 (151-172 microM) were the highest. The physiological significance of these results is discussed, especially in regard to the relative resistance of hBNP-32 to attack and the ability of the C-receptor ligands to compete with natriuretic peptides for hydrolysis by E-24.11.

scholarly journals Respective roles of kallikrein and endopeptidase 24.11 in the metabolic pathway of atrial natriuretic peptide in the rat

1990 ◽  
Vol 269 (3) ◽  
pp. 801-806 ◽  
Author(s):  
Y Vanneste ◽  
S Pauwels ◽  
L Lambotte ◽  
A Michel ◽  
R Dimaline ◽  
...  
Keyword(s):  
Atrial Natriuretic Peptide ◽  
Natriuretic Peptide ◽  
Cleavage Product ◽  
Constant Infusion ◽  
Maximal Inhibition ◽  
Blood Samples ◽  
Endopeptidase 24.11 ◽  
Rapid Clearance ◽  
Atrial Natriuretic

The metabolism of atrial natriuretic peptide (ANP) and Cys-105-Phe-106-cleaved ANP (ANP) was studied during constant infusion of 125I-labelled peptides in rats. Analysis of circulating radioactivity indicated rapid clearance of ANP and ANP′, with mean half-lives of 0.42 and 1.04 min respectively. H.p.l.c. fractionation of plasma taken during the infusion of labelled ANP revealed the presence of three radioactive fragments, the major one co-eluting with 125I-ANP′. These fragments correspond to cleavage products previously found to be generated in vitro by the action of endopeptidase 24.11 (E-24.11). On evaluating the effects of peptidase inhibitors, a significant increase in the half-life of ANP was observed with phosphoramidon (t1/2 7.8 min) and aprotinin (t1/2 5.4 min). A maximal inhibition of ANP degradation was obtained when both inhibitors were given simultaneously (t1/2 15 min). In blood samples taken during infusion of 125I-ANP and phosphoramidon, the intact peptide accounted for more than 90% of total circulating radioactivity, and no cleavage product was present in detectable amounts. Phosphoramidon had no effect on the metabolism of infused ANP′. In contrast, when 125I-ANP′ was infused together with aprotinin, the rate of degradation of the infused peptide was reduced by more than 80%. It is proposed that two different peptidase activities, E-24.11 and a kallikrein-like proteinase, are responsible for the cleavage of ANP in the circulation. The Cys-Phe-cleaved ANP would in turn be degraded by kallikrein and not by E-24.11.

scholarly journals Biological Activity of β-Human Atrial Natriuretic Peptide (β-hANP)

1987 ◽  
Vol 43 ◽  
pp. 91
Author(s):  
Yujiro Hayashi ◽  
Mayumi Furuya ◽  
Fuyuki Iwasa ◽  
Norio Ohnuma ◽  
Tamayo Hatoh ◽  
...  
Keyword(s):  
Biological Activity ◽  
Atrial Natriuretic Peptide ◽  
Natriuretic Peptide ◽  
Human Atrial Natriuretic Peptide ◽  
Atrial Natriuretic
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